Of the five major classes of immunoglobulin found in humans, immunoglobulin A (IgA) or secretory immunoglobulin generally is found in serous or mucus fluids Levels of IgA in serum are lower than that of IgG and about twice that of IgM. IgA levels are considerable in the sero-mucous secretion such as saliva, tears, nasal fluids, sweat, colostrum and secretions of the lungs and gastrointestinal tract. It is believed that IgA plays a major role in protecting the exposed epithelium.
It is not uncommon to find IgA in the form of a dimer, joined by a secretory protein which is synthesized by local epithelial cells. Dimers generally comprise antibodies with the same specificity. There are at least two IgA subclasses.
As with the other classes of antibodies, deficiency of IgA can occur transiently, for example in infants when maternal IgA levels wane when breast feeding is discontinued, or permanently, as in the not uncommon occurrence of patients with congenital IgA deficiency. Secretory IgA is known to have a beneficial protective effect on the intestinal mucosa in infants. Stoliar et al., Lancet 1976; i:1258; Williams & Gibbons, Science 1972; 177:697.
Eibl et al. (NEJM 1988; 319:1) found that oral feeding of an IgA-IgG preparation minimized the risk of infants not fed breast milk of contracting necrotizing enterocolitis. The immunoglobulin preparation was made from human serum, Cohn's Fraction II, by ion exchange chromatography. The preparations contained anywhere from 66% to 85% IgA, 15% to 34% IgG and 0.1% to 2% IgM.
Because of the various immunoglobulin deficiencies and the perceived benefits of passively immunizing patients with immunoglobulin deficiency, it is desirable to obtain preparations rich in immunoglobulin, and in particular IgA, for therapeutic uses.
U.S. Pat. No. 4,396,608 teaches a method of preparing an immune serum globulin (IgG) preparation suitable for injection. The method relates generally to preparing IgG rich compositions. A suitable starting material for the process of preparing the IgG rich composition is Cohn's Fraction II or Fraction II plus III. The starting wet paste or powder dissolved in water or physiologic solution is subjected to an acid treatment of about pH 3.5 to 5.0 and thereafter its ionic strength is reduced. All steps are performed at a temperature of 0.degree.-20.degree. C. Protein concentration is adjusted by conventional techniques such as ultrafiltration.
U.S. Pat. No. 4,477,432 relates to an immunoglobulin preparation containing not less than 70% IgG, suitable for oral administration. The immunoglobulin preparation has a pH of about 4-8, is sterile filtered and has a protein concentration of between 5-20%.
U.S. Pat. No. 4,499,073 relates to an immunoglobulin (IgG) preparation prepared by acid treatment wherein the monomer content is greater than about 90%. The starting paste or powder is dissolved in water or physiologic equivalent, the solution is adjusted to a pH of 3.5-5 and the ionic strength is adjusted to a low value.
In view of the apparent advantages of IgA rich preparations for therapeutic purposes it is desirable to have a procedure for obtaining IgA in sufficient quantity, of sufficient purity and suitable for human therapeutic purposes. The above patent references teach methods to obtain immunoglobulin rich preparations in general and those preparations are not necessarily rich in IgA.
Eibl et al., supra, relates to a chromatographic method for preparing an IgA rich preparation.
Skvaril & Brummelova (Coll. Czech. Chem. Comm. 1965; 30:2886) described a method of purifying IgA from the ethanol fraction III of placental serum using zinc salt in combination with ammonium sulphate and gel filtration. Thus, Cohn's Fraction III suspended in cold water was exposed to alumina gel and the eluted protein then was exposed to ammonium sulphate at 40-60% saturation. That cut was removed and dialyzed against water. The remaining proteins were dissolved in acetate buffer. The pass-through from a DEAE-Sephadex separation was obtained and zinc acetate was added to the solution. The precipitate that formed at 4.degree. C. was separated and the supernatant was treated with ammonium sulfate to a concentration of 2.05M. The precipitate was dissolved in water and upon passage through a Sephadex G200 column, fractions containing essentially pure IgA were identified. In vivo studies revealed that the IgA preparations contained trace amounts of IgG but no IgM was noted.
Anderson et al. (J Imm. 1970; 105:146) relates to a one-step isolation of IgA from small volumes of human serum using a bromoacetyl cellulose anti-IgA immunoabsorbent in a batch process. IgA was dissociated from the matrix with acetic acid and then dialyzed against phosphate buffer. The immunoabsorbent, however, is difficult to prepare. The preparations contained from 86% to 98.5% IgA, 0.7% to 11.2% IgG and 0.7% to 5.6% IgM.
Pejaudier et al., (Vox Sang. 1972; 23:165) obtained IgA from Cohn's Fraction III. Several purification schemes were tested. The preferred method comprised extraction of Fraction III with water at pH 5.6, precipitation with caprylic acid in acetate buffer and passage of the supernatant over a DEAE-cellulose matrix with an acetate buffer for elution. The preparations contained trace amounts of IgG and no IgM.
Kondoh et al. (Mol. Imm. 1987; 24:1219) described a procedure for the isolation of human secretory IgA using jacalin lectin. The jacalin was coupled with Sepharose 4B to produce an affinity column. Secreted IgA was obtained from human colostrum. The preparations contained trace amounts of IgM and no IgG.
Thus, it remains desirable to provide a simple procedure for purifying immunoglobulin A in sufficient quantities to produce compositions suitable for therapeutic uses.